Genome editing with Zinc Finger nucleases and TALENs has emerged as a promising technology for inducing localized sequence mutations in targeted genes. As CCR5 is the principal co-receptor for HIV, studies are underway to disrupt the gene in T cells and CD34+ Hematopoietic stem cells to protect the cells or their progeny from viral infection. So far, viral and plasmid vectors have been used for delivery of ZFNs/TALENs. However, this results in long-term expression of the nucleases, which may be potentially harmful, as increasing number of studies have documented dose-dependent toxicity of many engineered ZFN, presumably due to the generation of unintended double-stranded breaks at off-target sites. Transient expression of ZFNs/TALENs from mRNA may provide a feasible alternative as short term exposure to the nucleases is sufficient to cause permanent modification of the targeted gene. In this proposal, ZFN or TALEN expression from mRNA will be used as a strategy to disrupt CCR5 in CD34+ hematopoietic stem cells (HSCs). To overcome the hurdle of ZFN/TALEN mRNA delivery to CD34+ HSCs, which are difficult cells to transfect by conventional methods, we will develop lipid nanoparticles displaying a previously described DNA aptamer sgc8 that binds to Protein tyrosine kinase 7, expressed on the cells. Humanized BLT mice will be reconstituted with the ZFN/TALEN-modified CD34+ HSPCs and tested for protection from HIV challenge. Knowledge gained from the studies could lead to the development of a novel and safe approach to harness ZFN/TALEN for HIV gene therapy and even provide a potential method for direct delivery of the molecules to mobilized stem cells in vivo.